Electronic Measurements of Single-Molecule Processing by DNA Polymerase I (Klenow Fragment).
Bioconjugating single molecules of the Klenow fragment of DNA polymerase I into electronic nanocircuits allowed electrical recordings of enzymatic function and dynamic variability with the resolution of individual nucleotide incorporation events. Continuous recordings of DNA polymerase processing mu...
| Publicado en: | Journal of the American Chemical Society Vol. 135; no. 21; pp. 7855 - 7861 |
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| Autores principales: | , , , , , , , , |
| Formato: | Artículo |
| Publicado: |
American Chemical Society
5/29/2013
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| Materias: | |
| Acceso en línea: | Ver este registro en EBSCOhost |
| fields | @attributes: recordID: 1 pdfLink: plink: https://search.ebscohost.com/login.aspx?direct=true&db=hlh&AN=88117876&site=ehost-live header: @attributes: shortDbName: hlh uiTerm: 88117876 longDbName: Humanities International Complete uiTag: AN controlInfo: bkinfo: jinfo: jid: 00027863 ACS jtl: Journal of the American Chemical Society issn: 00027863 maglogo: N pubinfo: dt: 5/29/2013 vid: 135 iid: 21 pid: 997 pub: American Chemical Society artinfo: ui: 88117876 10.1021/ja311603r ppf: 7855 ppct: 6 formats: tig: atl: Electronic Measurements of Single-Molecule Processing by DNA Polymerase I (Klenow Fragment). aug: au: Olsen, Tivoli J. Yongki Choi Sims, Patrick C. Tolga Gul, O. Corso, Brad L. Chengjun Dong Brown, William A. Collins, Philip G. Weiss, Gregory A. affil: Department of Chemistry, University of California, Irvine, California 92697, United States Department of Physics and Astronomy, University of California, Irvine, California 92697, United States Department of Molecular Biology, University of California, Irvine, California 92697, United States su: DNA polymerases kinetics Single molecule research Biochemical templates Bioconjugates Electronic measurements Nucleotides Phosphodiesters sug: subj: DNA polymerases kinetics Single molecule research Biochemical templates Bioconjugates Electronic measurements Nucleotides Phosphodiesters ab: Bioconjugating single molecules of the Klenow fragment of DNA polymerase I into electronic nanocircuits allowed electrical recordings of enzymatic function and dynamic variability with the resolution of individual nucleotide incorporation events. Continuous recordings of DNA polymerase processing multiple homopolymeric DNA templates extended over 600 s and through >10 000 bond-forming events. An enzymatic processivity of 42 nucleotides for a template of the same length was directly observed. Statistical analysis determined key kinetic parameters for the enzyme's open and closed conformations. Consistent with these nanocircuit-based observations, the enzyme's closed complex forms a phosphodiester bond in a highly efficient process >99.8% of the time, with a mean duration of only 0.3 ms for all four dNTPs. The rate-limiting step for catalysis occurs during the enzyme's open state, but with a nearly 2-fold longer duration for dATP or dTTP incorporation than for dCTP or dGTP into complementary, homopolymeric DNA templates. Taken together, the results provide a wealth of new information complementing prior work on the mechanism and dynamics of DNA polymerase I. pubtype: Academic Journal doctype: Article src: R language: English refInfo: copyright: @attributes: flag: Y dt: @attributes: year: 2013 holdings: @attributes: islocal: N |
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